Introduction of genetic material into cells and tissues for controlling gene expression has significantly impacted research involving gene pathways and function, and provides promise for therapeutic application. The genetic level approach has inherent specificity not available with the vast majority of drugs. siRNAs hold great promise as potential therapeutic tools and are currently in clinical trials, targeting a wide range of clinical problems including cancer. Gene silencing is much more cost-effective, and leads to down-regulation of protein expression and function with greater potential specificity than small molecule inhibitors. In particular, siRNA treatment may target a single point mutation in a gene, while small molecule therapy to date does not precisely distinguish between mutant and normal gene products. Given the ability to determine specific gene alterations in each melanoma through identification of hotspot mutations, direct gene sequencing, or assays for gene amplification, each melanoma can be assigned a specific genetic signature. Although the siRNA may be taken up by many cells, only cells with a mutated gene or activated signaling protein are affected by targeted gene therapy, thereby allowing normalization of pathway signaling in melanomas without adversely affecting normal cells.
As with delivery of many proteins, degradation of nucleic acids and poor bioavailability from the gastrointestinal tract are major hurdles to the oral delivery of siRNAs. Even with intravenous delivery, conventional siRNA is rapidly degraded by serum factors and does not reach its targets. Topical application of nucleic acids offers great therapeutic advantages, both for suppressing genes in lesional skin (for example and without limitation, to treat metastases in skin) and for transdermal delivery to internal targets. Application is painless and easily controlled, and skin is highly accessible. The effective physical barrier in the epidermis is localized mainly to the outermost area of epidermis, the stratum corneum, and to a lesser extent the deeper epidermis. This epidermal barrier protects against extensive water loss (inside-out) and against the entry of environmental substances (outside-in), including nucleic acids. Mechanical approaches, such as ultrasound, laser and injection, have been used to facilitate penetration through the mouse stratum corneum and drive siRNA into skin, but require specialized equipment, limit the area of delivery, and potentially harm the skin. These challenges emphasize the need for an easily applied transdermal system for delivering suppressive nucleic acids that is able to transit the stratum corneum.
Direct targeting of a skin disorder is an ideal model for gene suppressive therapy. However, the commercially available materials to suppress genes in vitro have been marginally successful, at best, for delivering genetic material into primary cultured cells, such as keratinocytes (KCs) and melanocytes. Furthermore, the outer layers of skin function as an anatomic barrier that traditionally prevents the penetration of nucleic acids and proteins into skin and, from dermis, into the circulation [Prausnitz et al., Nat Biotechnol 26: 1261-1268 (2008)]. Thus, traversing this layer to transfer sufficient amounts of oligonucleotides has been a challenge.
The skin is the largest organ of the body and contains three layers: the epidermis, dermis, and subcutaneous tissue. The epidermis is the outer layer of skin. The thickness of the epidermis varies in different types of skin. It is the thinnest on the eyelids at 0.05 mm and the thickest on the palms and soles at 1.5 mm. The epidermis contains 4 major layers of progressively more differentiated cells. From bottom to top the layers are named:                stratum basale        stratum spinosum        stratum granulosum        stratum corneum        
The bottom layer, the stratum basale, has cells that are shaped like columns. In this layer the cells divide and push already-formed cells into higher layers. As the cells move into the higher layers, they flatten, become more mature and eventually “die” and are shed. The top layer of the epidermis, the stratum corneum, is made of flattened skin cells that are shed; it takes about 4 weeks from cells of the stratum basale to reach the stratum corneum and subsequently be shed.